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Figure 3 SMYD3 knockdown increases mesendodermal marker expression in differentiating mESCs. (A,B) mRNA levels of mesendodermal markers (Eomes, Mesp1, Sox17, Foxa2, T/Brachyury) were measured by qRT-PCR. Data were normalized to Gapdh. Means and SEM from three independent experiments are shown. Significance was calculated by 2-way Anova, followed by Bonferroni post hoc test. * p < 0.05, *** p < 0.001 value for the significance is shown in the plot. (C) eGFP reporter fluorescence was analyzed in EBs infected with Sh-Scramble/Brachyury-eGFP and Sh-SMYD3/Brachyury-eGFP at day 6 of differentiation. Scale bar: 20 μm. (D) Primitive ectoderm genes (Otx2, Fgf5, Sox1) expression was measured by qRT-PCR at different time points. Means and SEM from three independent experiments are shown.